The goal is to define the molecular mechanism of calcium transport across the small intestine, and to provide a molecular understanding of the effects of vitamin D in the control of calcium transport. The methods to be used involve radioactive tracer and spectrophotometric techniques to measure the fluxes across the intact intestinal epithelium, across isolated brush border and basolateral plasma membranes, and reconstituted phospholipid liposomes. Towards the isolation, purification and characterization of the peptides responsible for active calcium transport, we will identify the peptides of the pure basolateral membrances using gel electrophoresis, histochemical staining, 32P-phosphorylaton and general peptide labeling techniques. Upon identification of Ca-ATPase we will attempt to isolate the peptide and incorporate it into phospholipid membranes in order to study the molecular basis of active calcium trnasport. Identification of the calcium transporting enzyme in the epithelial plasma membranes will further permit us to study the molecular responses to the active form of vitamin D, i.e. 1,25(OH)2D3. In addition, we plan to study the general trnasport characteristics of the pure, isolated plasma membranes.